Different stability of N- and C-domain of diferric ovotransferrin in urea and application to the determination of iron distribution between the two domains

The study of guanidine-HCl or thermal denaturation of diferric ovotransferrin (Fe2Tf) has revealed a simultaneous unfolding of the two domains of the protein (Ikeda et al. (1985) FEBS Lett. 182, 305-309). In urea denaturation of Fe2Tf, however, two distinct steps of unfolding were observed in the urea concentration range from 4.5 to 9 M at pH 8.0 and 37 degrees C by measuring the residual iron-bound protein (absorbance at 465 nm) and the remaining folded structures (circular dichroism at 222 nm). From a study of urea denaturation of partially iron-saturated Tf whose iron preferentially occupied the N-domain, it was found that the first and the second steps of denaturation corresponded to those of the N-terminal (4.5-6 M urea) and C-terminal domains (over 7 M urea), respectively. The N-domain of Fe2Tf was selectively unfolded in 7 M urea and digested with trypsin to provide an iron-bound C-terminal fragment (42 kDa) in good yield (about 80% of theoretical). The kinetic analysis of the decrease in A465 of Fe2Tf in 9 M urea showed that the N-domain unfolded 3 x 10(2) times faster than the C-domain. With partially iron-saturated Tf, the decrease of A465 in 9 M urea also proceeded in a biphasic manner and the ratio, the decrement in A465 of the rapid phase/the decrement in A465 of the slow phase, gave the value of iron distribution as Fe at the N-site/Fe at the C-site.     

Effect of physical training on immunoreactive gamma-glutamyltransferase in human plasma

The effects of 10-week physical training on both the activity and concentration of gamma-glutamyltransferase (gamma-GT) in plasma were investigated on 7 sedentary healthy male students. The training consisted of running over 5 km, 6 times/week. The maximal oxygen uptake (Vo2max) and 12-min field performance increased significantly after training, from 43.5 and 2,683.7 to 48.1 ml.kg-1.min-1 and 2,931.1 m, respectively. After training the resting plasma gamma-GT concentration decreased to half, whereas the gamma-GT activity did not vary substantially. The training did not affect the response to the Vo2max test of either concentration or activity of gamma-GT. These results may suggest that plasma gamma-GT concentration is useful as an index of the extent of physical training.     

Purification and characterization of chitinase produced by Streptomyces erythraeus

A chitinase was purified from the culture filtrate of Streptomyces erythraeus (SE). The enzyme (SE chitinase) has a molecular weight of 30,000 and pI 3.7, and shows optimal activity at pH 5.0 with an optimal ionic strength of less than 0.2 M NaCl. SE chitinase could hydrolyze chitin and its derivatives, but could not hydrolyze cell walls of Micrococcus lysodeikticus. The substrate specificity of SE chitinase was compared with those of hen egg white (HEW) and SE lysozymes. The binding mode of the chitinase to substrates was investigated using chitooligosaccharides and their derivatives. The results showed that the binding mode of SE chitinase to the substrate is similar to that of HEW and SE lysozymes.     

Characterization of very low density lipoprotein from Watanabe heritable hyperlipidemic rabbits

We investigated the properties of very low density lipoprotein (VLDL) from two types of Watanabe heritable hyperlipidemic (WHHL) rabbits: one with a high incidence of coronary atherosclerosis (type 1), and the other with a low incidence (type 2). When incubated with mouse peritoneal macrophages, VLDL from type 1 WHHL rabbit (type 1-VLDL) stimulated cholesteryl ester synthesis 10.5-fold more than VLDL from the type 2 WHHL rabbit (type 2-VLDL) did. Moreover, a similar difference was seen in the stimulation of cholesteryl ester synthesis in peritoneal macrophages isolated from the WHHL rabbits. The mass ratios of cholesterol to protein in type 1- and type 2-VLDL were 5.69 and 2.05, respectively. Agarose gel electrophoresis of type 1-VLDL showed beta mobility, and that of type 2-VLDL showed pre-beta mobility. No difference was seen between the sizes of VLDL particles of the two types. The amount of apolipoprotein E in type 1-VLDL was greater than that in type 2-VLDL. In conclusion, the difference between type 1 and type 2 WHHL rabbits is at least partly due to the presence in type 1 animals of VLDL particles rich in cholesteryl esters and apolipoprotein E, particles which are very similar to beta-VLDL in conformation.     

Litter-fall and reproductive seasonalities in aLeucaena leucocephala forest at Chichijima,Ogasawara (Bonin) Islands

Seasonal changes in litter components were measured in a closedLeucaena leucocephala forest stand in Chichijima, Ogasawara (Bonin) Islands, situated in a wet subtropical climate. The phytomass of the stand was estimated as 5.62, 1.47 and 0.448 kg d.w. m⁻² for stems, branches and leaves, respectively. Litter from the canopy was collected semimonthly by ten 0.5 m² traps. Leaf-litter production rate was low in spring, medium in summer and autumn, and peaked in mid-winter. Annual production of leaf-litter was 498 g d.w. m⁻². Annual turnover rate of canopy leaves was 1.1. Flower-litter showed biannual production with peaks in spring and autumn, and the former peak largely exceeded the latter one in quantity. Seed maturation needed about 3 months. Annual production of reproductive organs was 89.2, 89.4, 19.1 and 99.4 g d.w. m⁻² for flowers, mature seeds, immature ones and pods, respectively. Number, of mature seeds disseminated exceeded 2,500 m⁻². Annual production of branch- and bark-litter was 345 g d.w.m⁻². Annual amount of plant litter from the canopy was as large as 1,150 g d.w. m⁻², showing higher productivity and higher rate of matter turnover in this forest than in temperate forests.     

Sexual agglutinin of mating-type minus gametes inChlamydomonas reinhardtii

The sexual agglutinin from the mating-type minus gametes ofChlamydomonas reinhardtii was purified by gel filtration and hydroxyapatite chromotography. The minus agglutinin was identified as a single glycopolypeptide termed Agg(-) of very high molecular weight by SDS-poly-acrylamide gel electrophoresis. It was also observed as a glycoprotein with agglutinin activity on non-SDS polyacrylamide gels. The native agglutinin appeared to be composed of a complex of Agg(-) subunits. It consisted of about 60% protein and 40% carbohydrate and the activity was diminished by a mild periodate oxidation. When the plus agglutinin was also purified and compared with the minus agglutinin, it was found that both agglutinins migrate in the same position by SDS-polyacrylamide gel electrophoresis, whereas their behaviors on gel filtration and hydroxyapatite chromatography are different.Abbreviations mt ^+/– mating-tape plus or minus - SDS sodium dodecyl sulfate - V_e elution volume - V_o void volume - kDa kilodalton     

Differential regulation of high-affinity agonist binding to muscarinic sites in the rat heart, cerebellum, and cerebral cortex

The muscarinic agonist [3H]cismethyldioxolane ([3H]CD) was used to characterize the effects of regulators upon high-affinity agonist binding sites of the rat heart, cerebral cortex and cerebellum. Comparative studies with sodium ions (Na+), magnesium ions (Mg++), N-ethylmaleimide (NEM) and the guanine nucleotide Gpp(NH)p revealed tissue-specific effects. Mg++ preferentially enhanced while Gpp(NH)p and NEM reduced high-affinity [3H]CD binding in the heart and cerebellum. By comparison NEM enhanced high-affinity agonist binding in the cerebral cortex while Gpp(NH)p and Mg++ had little or no effect. Kinetic studies support an allosteric mechanism for these effects and provide further evidence for muscarinic receptor subtypes in mammalian tissues.     

Alternation in colonization behaviors of Escherichia coli cells with rpoS or yggE deficiency on solid surfaces

Colonization on a solid surface is influenced by the cell surface appendages such as flagella and curli, of which expressions are regulated by rpoS gene encoding a sigma factor. In this study, we investigated the effect of rpoS or yggE (a rpoS‐related and stress‐responsive gene) deficiency on the colonization of Escherichia coli BW25113. Under a static condition, the deletion of rpoS or yggE induced 3.9‐ and 3.7‐fold higher colonization as compared to wild‐type cells, respectively, on the solid surfaces. However, under a liquid flow condition, only ΔyggE cells maintained the stable colonization on the surface, and the values of cell layer thickness and cell coverage on the surface were 17 and 9.2 times as high as those of wild‐type cells, respectively. Gene expression analyses revealed that the deletion of rpoS or yggE positively impacted the expressions of genes involved in flagellum formation. On the other hand, curli assembly was severely prohibited by the rpoS deficiency. Here, we proposed that the plentiful flagella on the ΔrpoS and ΔyggE cell surfaces facilitated mainly the colonization under the static condition. Meanwhile, curli existing on the ΔyggE cell surface played an important role in keeping stable cell attachment and developing attached colonies under the flow stress condition. Biotechnol. Bioeng. 2013; 110: 1050–1056. © 2013 Wiley Periodicals, Inc.     

Long-term changes in the population size and geographical distribution of Pennella sp. (Copepoda) on the saury,Cololabis saira,in the western North Pacific Ocean and adjacent seas

Changes in the population size and geographical distribution of the mesoparasitic copepod Pennella sp. (Siphonostomatoida: Pennellidae) on the saury, Cololabis saira, were studied for a period of six years (1981–86) in the western North Pacific Ocean and adjacent seas. The parasite was first recorded in the western North Pacific in 1981. Its population size increased rapidly in 1982–83 and declined slightly in 1984. During these years, infected fish were widely distributed in the western and central North Pacific and also found in the Sea of Okhotsk and the Sea of Japan. However, the population declined dramatically in 1985 and its distributional range was reduced. The parasite disappeared in 1986. The observed frequency distributions of parasites on the host population were the Poisson in 1981 and 1985, but those during 1982–1984 were over-dispersed and fitted the negative binomial.Parasites of the saury, Cololabis saira (Brevoort)-IV.